Recent studies have demonstrated the feasibility of using ELISPOT assays to measure the antigen-specific responsiveness of individual T cells taken from patients. This approach is relatively simple, reproducible, and when necessary it can be modified to yield great sensitivity. Consequently, ELISPOT assays are already being used clinically in state of the art trials to quantitate T cell responses to tumor- and virus-specific vaccines. This approach, to date, has not been used systematically to monitor alloreactivity against minor transplantation antigens and tumor antigens in the setting of human hematopoietic stem cell transplantation. We are adapting published methods for measuring responses to defined exogenous antigens for use in monitoring T cell responses to allogeneic recipient cells. When appropriately standardized we will use the resulting assay for serial monitoring of the alloreactive activity of donor leukocytes collected before transplant. By comparing these assays with post-transplant clinical course and chimerism data, we hope to establish whether pretransplant assessment can be used in predicting the clinical course after transplantation. In addition, we will serially monitor the alloreactivity of T cells obtained post-transplant. Taken together these studies seek to identify ex vivo findings which may correlate with or even predict clinical findings such as donor hematopoietic and immune cell engraftment, graft-versus-host disease, and graft versus tumor effect.